Hydrogel-Encapsulated Beads Enable Proximity-Driven Encoded Library Synthesis and Screening

Area of Science:

• Biotechnology
• Chemical Biology
• Materials Science

Background:

• Encoded combinatorial libraries offer vast chemical diversity but are limited to affinity-based screening.
• Reformatting these libraries to "one-bead-one-compound" solid-phase formats is crucial for broader screening applications.
• Existing methods lack efficient ways to convert selection outputs for diverse screening modalities.

Purpose of the Study:

• To develop a method for reformatting DNA-encoded libraries into a "one-bead-one-compound" format using hydrogel-encapsulated magnetic beads.
• To demonstrate the compatibility of these beads with various chemical and biological transformations.
• To validate the reformatting approach using mRNA display libraries and assess enrichment efficiency.

Main Methods:

• Synthesized uniform magnetic microbeads encapsulated in polyacrylamide hydrogel shells via emulsion polymerization.
• Functionalized the hydrogel beads with amine, alkyne, and oligonucleotide moieties.
• Performed in-gel transformations including acylation and enzymatic DNA ligation.
• Utilized transcription, hybridization, and puromycin labeling for mRNA display library reformatting.
• Employed fluorescence-activated cell sorting (FACS) for bead enrichment and screening.

Main Results:

• Achieved uniform hydrogel-encapsulated magnetic beads (7 ± 2 μm) compatible with diverse functionalization.
• Successfully reformatted mRNA display libraries, demonstrating colocalization of RNA synthesis and translation.
• Enriched two control epitope templates (V5, HA) with significant fold-increases (50- and 99-fold) from a library screen.
• Demonstrated the utility of proximity-driven library synthesis and magnetic manipulation for library reformatting.

Conclusions:

• Hydrogel-encapsulated magnetic beads provide a versatile platform for reformatting encoded combinatorial libraries.
• This technology unlocks activity-based and cellular screening capabilities for previously inaccessible library formats.
• The developed method offers a scalable and efficient approach for transforming encoded libraries for broader drug discovery efforts.